Neurotransmission requires a precise number and arrangement of receptors, ion channels, and adhesion molecules at synapses. Alteration of the localization or levels of these proteins at the postsynaptic membrane regulates synapse function, thereby strengthening or weakening synaptic connections in the brain. In all eukaryotic cells, removal of diverse surface membrane proteins occurs by clathrin-mediated endocytosis. Although previous studies have helped define the endocytic machinery in nonneuronal cells and in the presynaptic nerve terminal, the location and regulation of clathrin- mediated endocytosis within postsynaptic compartments and its functional role in synaptic signaling remain poorly understood. To address these important questions, my laboratory has initiated a program of cell biological and physiological studies to analyze the endocytic machinery of dendritic spines - the primary postsynaptic compartment in the mammalian brain. We have found that dendritic spines contain a zone of clathrin assembly and endocytosis adjacent to, but spatially segregated from, the postsynaptic density. This `endocytic zone'forms and persists over long periods of time, and it serves to concentrate cargo destined for internalization. Further, we have recently identified a direct physical link between the postsynaptic scaffold complex and clathrin endocytic machinery and found a novel requirement for localized endocytosis and recycling in maintaining a synaptically proximate pool of glutamate receptors. Taking advantage of these preliminary data and our ability to monitor and manipulate endocytosis in dendritic spines, we propose to define the underlying molecular and cellular mechanisms that form, maintain, and regulate the endocytic zone of spines, and determine the functional role of such zones in synapse maintenance and modification. This work will provide insight into fundamental mechanisms that underlie synapse formation and synaptic plasticity. Moreover, because clathrin-mediated endocytosis regulates neuronal responsiveness to a wide range of pathologic insults and therapeutic agents relevant to numerous neurologic and psychiatric diseases these studies hold promise for the development of novel therapeutic strategies. PUBLIC HEALTH RELEVANCE: The proposed research will uncover molecular mechanisms that regulate brain cell communication at synapses. Abnormal function of synapses contributes to epilepsy, memory decline, depression, autism, schizophrenia, and addiction. By helping to understand how nerve cell synapses are adjusted during brain development and modified as we learn, the proposed research will define novel targets and therapeutic strategies for these devastating neurological and psychiatric disorders, which currently have a profound negative impact on public health.